This research is directed towards the genetic and biochemical characterization of early steps in lambda phage DNA replication. We are attempting to identify both phage and host genes and their products that mediate these steps. Lambda DNA circles in lysogens are rapidly nicked by two processes and also become associated with what appears to be the host membrane. We have evidence that one type of nick is associated with the formation of rapidly sedimenting complexes that contain bacterial membranes. Membrane-associated nicking in vivo is not strand specific, but we are still trying to determine if it is site specific. Failure to detect phage structural gene(s) responsible for this type of nicking or membrane association has led us to examine E. coli mutants defective in DNA replication for their ability to mediate formation of lambda DNA-host rapidly sedimenting complexes along with concomitant membrane-associated nicking. We are expanding this investigation to new dnats mutants and to other mutations in E. coli that are known to affect lambda DNA replication.